SecYEG-mediated translocation in a model synthetic cell

Area of Science:

• Synthetic biology
• Biochemistry
• Membrane protein biophysics

Background:

• Giant unilamellar vesicles (GUVs) are crucial model systems for synthetic cell research.
• Incorporating functional membrane proteins into GUVs remains a significant challenge for creating advanced synthetic cells.
• Key membrane proteins are needed for essential cellular functions like transport, energy conversion, and replication.

Purpose of the Study:

• To develop a method for incorporating the bacterial translocase SecYEG into GUVs.
• To create functionalized GUVs (SecGUVs) suitable for large-scale synthetic biology applications.
• To demonstrate that SecYEG retains its translocation activity within the GUVs.

Main Methods:

• Reconstitution of the SecYEG protein complex into small unilamellar vesicles (SUVs).
• Fusion of SecYEG-containing SUVs with pre-formed GUVs to generate SecGUVs.
• Assessment of SecYEG translocation efficiency within the engineered GUVs.

Main Results:

• Successfully produced SecGUVs by fusing SecYEG-reconstituted SUVs with GUVs.
• Maintained a high protein:lipid ratio in the engineered SecGUVs.
• Demonstrated that SecYEG translocation efficiency is not compromised by its incorporation into GUVs.

Conclusions:

• The developed method enables the creation of robust, functional proteo-GUVs.
• SecGUVs are versatile compartments for advancing synthetic cell formation and growth.
• This work provides a foundation for engineering more complex synthetic cellular systems.